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Recently, in situ YSI EXO2 phycocyanin fluorescence probes have been widely deployed as a means to determine cyanobacterial abundance in drinking water sources, yet few studies have evaluated the effects of natural organic matter (NOM) and the ambient water temperature on the probe readings. In this study, Suwannee River NOM was added to laboratory cultivated cyanobacterial species to test the performance of the phycocyanin probe. The impact of temperature on phycocyanin fluorescence was evaluated by monitoring the laboratory cultivated cyanobacterial species and extracted phycocyanin pigment. Additionally, in situ phycocyanin fluorescence of the field samples from the water intake of a drinking water treatment plant (DWTP) in 2018 were compared with grab sample laboratory taxonomic analyses. We found: (1) the presence of Suwannee River NOM leads to the decrease in cell-bound cyanobacterial phycocyanin readings; (2) increasing ambient water temperature reduces dissolved and cell-bound cyanobacterial phycocyanin readings; (3) field study phycocyanin probe readings significantly correlated with the total cyanobacterial biovolume (R = 0.73, p < 0.1), and the relationship depends on the biovolume of dominant cyanobacterial species; (4) phycocyanin probe readings have a strong positive correlation with the natural light intensities; and (5) probe users should be fully aware of the sources of interferences when interpreting the results and apply the other physical-chemical parameters data simultaneously generated by the fluorometry to improve the probe’s measurements.

Health-related concerns about cyanobacteria-laden sludge of drinking water treatment plants (DWTPs) have been raised in the past few years. Microscopic taxonomy, shotgun metagenomic sequencing, and microcystin (MC) measurement were applied to study the fate of cyanobacteria and cyanotoxins after controlled sludge storage (stagnation) in the dark in a full-scale drinking water treatment plant within 7 to 38 days. For four out of eight dates, cyanobacterial cell growth was observed by total taxonomic cell counts during sludge stagnation. The highest observed cell growth was 96% after 16 days of stagnation. Cell growth was dominated by potential MC producers such as Microcystis, Aphanocapsa, Chroococcus, and Dolichospermum. Shotgun metagenomic sequencing unveiled that stagnation stress shifts the cyanobacterial communities from the stress-sensitive Nostocales (e.g., Dolichospermum) order towards less compromised orders and potential MC producers such as Chroococcales (e.g., Microcystis) and Synechococcales (e.g., Synechococcus). The relative increase of cyanotoxin producers presents a health challenge when the supernatant of the stored sludge is recycled to the head of the DWTP or discharged into the source. These findings emphasize the importance of a strategy to manage cyanobacteria-laden sludge and suggest practical approaches should be adopted to control health/environmental impacts of cyanobacteria and cyanotoxins in sludge.

The excessive proliferation of cyanobacteria in surface waters is a widespread problem worldwide, leading to the contamination of drinking water sources. Short- and long-term solutions for managing cyanobacterial blooms are needed for drinking water supplies. The goal of this research was to investigate the cyanobacteria community composition using shotgun metagenomics in a short term, in situ mesocosm experiment of two lakes following their coagulation with ferric sulfate (Fe2(SO4)3) as an option for source water treatment. Among the nutrient paramenters, dissolved nitrogen was related to Microcystis in both Missisquoi Bay and Petit Lac St. François, while the presence of Synechococcus was related to total nitrogen, dissolved nitrogen, dissolved organic carbon, and dissolved phosphorus. Results from the shotgun metagenomic sequencing showed that Dolichospermum and Microcystis were the dominant genera in all of the mesocosms in the beginning of the sampling period in Missisquoi Bay and Petit Lac St. François, respectively. Potentially toxigenic genera such as Microcystis were correlated with intracellular microcystin concentrations. A principal component analysis showed that there was a change of the cyanobacterial composition at the genus level in the mesocosms after two days, which varied across the studied sites and sampling time. The cyanobacterial community richness and diversity did not change significantly after its coagulation by Fe2(SO4)3 in all of the mesocosms at either site. The use of Fe2(SO4)3 for an onsite source water treatment should consider its impact on cyanobacterial community structure and the reduction of toxin concentrations.

Freshwater bodies and, consequently, drinking water treatment plants (DWTPs) sources are increasingly facing toxic cyanobacterial blooms. Even though conventional treatment processes including coagulation, flocculation, sedimentation, and filtration can control cyanobacteria and cell-bound cyanotoxins, these processes may encounter challenges such as inefficient removal of dissolved metabolites and cyanobacterial cell breakthrough. Furthermore, conventional treatment processes may lead to the accumulation of cyanobacteria cells and cyanotoxins in sludge. Pre-oxidation can enhance coagulation efficiency as it provides the first barrier against cyanobacteria and cyanotoxins and it decreases cell accumulation in DWTP sludge. This critical review aims to: (i) evaluate the state of the science of cyanobacteria and cyanotoxin management throughout DWTPs, as well as their associated sludge, and (ii) develop a decision framework to manage cyanobacteria and cyanotoxins in DWTPs and sludge. The review identified that lab-cultured-based pre-oxidation studies may not represent the real bloom pre-oxidation efficacy. Moreover, the application of a common exposure unit CT (residual concentration × contact time) provides a proper understanding of cyanobacteria pre-oxidation efficiency. Recently, reported challenges on cyanobacterial survival and growth in sludge alongside the cell lysis and cyanotoxin release raised health and technical concerns with regards to sludge storage and sludge supernatant recycling to the head of DWTPs. According to the review, oxidation has not been identified as a feasible option to handle cyanobacterial-laden sludge due to low cell and cyanotoxin removal efficacy. Based on the reviewed literature, a decision framework is proposed to manage cyanobacteria and cyanotoxins and their associated sludge in DWTPs.

The impact of oxidation on mitigation of cyanobacteria and cyanotoxins in drinking water treatment sludge was investigated at the laboratory and treatment plant scales. Two common oxidants, KMnO4 (5 and 10 mg/L) and H2O2 (10 and 20 mg/L) were applied under controlled steady-state conditions. Non-oxidized and oxidized sludge was left to stagnate in the dark for 7 to 38 days. Controlled laboratory trials show that KMnO4 and H2O2 decreased cell counts up to 62% and 77%, respectively. The maximum total MC level reduction achieved after oxidation was 41% and 98% using 20 mg/L H2O2 and 10 mg/L KMnO4, respectively. Stagnation caused cell growth up to 2.6-fold in 8 out of 22 oxidized samples. Microcystin (MC) producer orders as Chroococcales and Synechococcales were persistent while Nostocales was sensitive to combined oxidation and stagnation stresses. In parallel, two on-site shock oxidation treatments were performed in the DWTP’s sludge holding tank using 10 mg/L KMnO4. On-site shock oxidation decreased taxonomic cell counts by up to 43% within 24 h. Stagnation preceded by on-site shock oxidation could increase total cell counts by up to 55% as compared to oxidation alone. The increase of cell counts and mcyD gene copy numbers during stagnation revealed the impact of oxidation/stagnation on cyanobacterial cell growth. These findings show the limitations of sludge oxidation as a strategy to manage cyanobacteria and cyanotoxins in sludge and suggest that alternative approaches to prevent the accumulation and mitigation of cyanobacteria in sludge should be considered.

ABSTRACT Wastewater-based epidemiology has emerged as a promising tool to monitor pathogens in a population, particularly when clinical diagnostic capacities become overwhelmed. During the ongoing COVID-19 pandemic caused by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), several jurisdictions have tracked viral concentrations in wastewater to inform public health authorities. While some studies have also sequenced SARS-CoV-2 genomes from wastewater, there have been relatively few direct comparisons between viral genetic diversity in wastewater and matched clinical samples from the same region and time period. Here we report sequencing and inference of SARS-CoV-2 mutations and variant lineages (including variants of concern) in 936 wastewater samples and thousands of matched clinical sequences collected between March 2020 and July 2021 in the cities of Montreal, Quebec City, and Laval, representing almost half the population of the Canadian province of Quebec. We benchmarked our sequencing and variant-calling methods on known viral genome sequences to establish thresholds for inferring variants in wastewater with confidence. We found that variant frequency estimates in wastewater and clinical samples are correlated over time in each city, with similar dates of first detection. Across all variant lineages, wastewater detection is more concordant with targeted outbreak sequencing than with semi-random clinical swab sampling. Most variants were first observed in clinical and outbreak data due to higher sequencing rate. However, wastewater sequencing is highly efficient, detecting more variants for a given sampling effort. This shows the potential for wastewater sequencing to provide useful public health data, especially at places or times when sufficient clinical sampling is infrequent or infeasible.

Wastewater surveillance for SARS-CoV-2 RNA is a relatively recent adaptation of long-standing wastewater surveillance for infectious and other harmful agents. Individuals infected with COVID-19 were found to shed SARS-CoV-2 in their faeces. Researchers around the world confirmed that SARS-CoV-2 RNA fragments could be detected and quantified in community wastewater. Canadian academic researchers, largely as volunteer initiatives, reported proof-of-concept by April 2020. National collaboration was initially facilitated by the Canadian Water Network. Many public health officials were initially skeptical about actionable information being provided by wastewater surveillance even though experience has shown that public health surveillance for a pandemic has no single, perfect approach. Rather, different approaches provide different insights, each with its own strengths and limitations. Public health science must triangulate among different forms of evidence to maximize understanding of what is happening or may be expected. Well-conceived, resourced, and implemented wastewater-based platforms can provide a cost-effective approach to support other conventional lines of evidence. Sustaining wastewater monitoring platforms for future surveillance of other disease targets and health states is a challenge. Canada can benefit from taking lessons learned from the COVID-19 pandemic to develop forward-looking interpretive frameworks and capacity to implement, adapt, and expand such public health surveillance capabilities.